STRUCTURED LIFE COURSE ANALYSIS OF DNA METHYLATION ASSOCIATIONS WITH INTERNALIZING BEHAVIOR AND ANXIETY FROM AGES 3-22

Mental health challenges often emerge early in development, with childhood and adolescence representing key periods of vulnerability. Internalizing symptoms, which include anxious and depressive symptoms, are among the most common psychological difficulties in youth and are typically expressed inwardly through withdrawal, sadness, and worry. Across several lines of research, including developmental psychopathology and molecular psychiatry, DNA methylation (DNAm) has often been conceptualized as a biologically embedded mechanism through which early-life experiences shape vulnerability to later psychopathology. Evidence from longitudinal data suggests that DNAm profiles shift in parallel with changes in psychological states, suggesting reciprocal and time-varying associations between symptoms and epigenetic regulation. These perspectives suggest that DNAm signatures associated with internalizing symptoms may reflect a combination of biological vulnerability, cumulative exposure, and contemporaneous symptom-related processes. In this study, we used the structured life course modeling approach (SLCMA) to examine time-sensitive associations between internalizing symptoms at multiple timepoints and epigenome-wide DNAm at both age 9 and 15 in a representative birth cohort from large U.S. cities (the Future of Families and Child Wellbeing Study, n=2,020). We applied a life-course modeling approach to evaluate the associations between DNAm and internalizing symptoms (measured using mother reported Child Behavioral Checklist) using two frameworks: (1) sensitive periods, the impact of internalizing symptoms are strongest at a specific timepoint (i.e., age 3, 5, 9, or 15), and (2) accumulation, where the impact of internalizing symptoms increases as symptoms accumulate over time. We also examine to what extent these profiles are predictive of later Anxiety (measured at age 22). At age 9, a total of 30 CpG sites were identified that passed FDR-q < 0.05 threshold, and most were associated with internalizing symptoms at age 9 (n = 25; 83.3%). At age 15, a total of 23 CpG sites were identified that passed FDR-q < 0.05 threshold, and most were associated with internalizing symptoms at age 9 (n = 20, 87%). Across both analyses, 14 CpG sites replicate between the two time points. Of the 14 CpG site hits, 8 CpG’s showed significant prospective associations of internalizing symptoms at 9 predicting later DNAm at 15. Another subset of 8 CpG’s showed an alternative association where internalizing symptoms may be an outcome of changes in DNAm. Five of the CpG sites that showed significant longitudinal associations showed bi-directional effects. These models suggest that these hits may have heterogeneous relationships with internalizing symptoms. Understanding the time-sensitive effects of early-life internalizing symptoms have on child and adolescent development through DNAm may provide insights into how these experiences become biologically embedded and lead to poorer health outcomes later in life.